Inhibition of the 5-lipoxygenase pathway utilizing certain 2,2&#39;-alkyldiyl bis(thio)bis-imidazoles and derivatives

ABSTRACT

Novel compounds, pharmaceutical compositions and a method of inhibiting the 5-lipoxygenase products in an animal in need thereof which comprises administering an effective, 5-lipoxygenase pathway inhibiting amount of a 2,2&#39;-[1,2-ethanediylbis-(thio)]-bis-1H-imidazole or 2,2&#39;-[1,3-propan-2-onediylbis-(thio)]bis-1H-imidazole, or a pharmaceutically acceptable salt thereof, to such animal.

CROSS REFERENCE TO RELATED APPLICATIONS

This is a divisional of application Ser. No. 856,735 filed Apr. 28, 1986now U.S. Pat. No. 4,728,656 which is a continuation-in-part of Ser. No.808,396 filed Dec. 12, 1985, now abandoned.

BACKGROUND OF THE INVENTION

This invention relates to novel compounds, pharmaceutical compositionsand a method of inhibiting the 5-lipoxygenase pathway of arachidonicacid metabolism in an animal in need thereof which comprisesadministering to such animal an effective, 5-lipoxygenase pathwayinhibiting amount of a 2,2-[1,2-ethanediyl-bis-(thio)bis-1H-imidazole, a2,2-[1,3-propan-2-onediylbis(thio)]bis-1H-imidazole or apharmaceutically acceptable salt thereof.

Hill, U.S. Pat. No. 4,188,397, issued Feb. 12, 1980 discloses compoundsof the formula ##STR1## wherein:

R is 4-monosubstituted phenyl wherein said substituent is selected frommethoxy, methylthio, trifluoromethyl, chloro, fluoro, bromo ormethylenedioxy when taken with an adjacent position on the phenyl ring;

R² and R³ are both H or one is H and the other is CH₃ ;

n is 0, 1 or 2; and

X is, among others, (CH₂)₂ and CH₂ C(O)CH₂. Hill also discloses thatsuch compounds are useful as antiarthritic agents, as confirmed by theirability to inhibit adjuvant induced polyarthritis in rats; and are alsouseful to regulate cell mediated immunity, as confirmed by theoxazolone-induced contact sensitivity test procedure which measureschanges in mouse paw edema produced by administration of test compounds.

The adjuvant-induced polyarthritis assay in rats is useful in detectingcompounds which are inhibitors of prostanoid synthesis, mediated by theprostanoids formed by the enzyme cyclooxygenase, but is of no knownutility in detecting or suggesting compounds which are inhibitors of thegeneration of 5-lipoxygenase products (such as HETES, LTB₄ andpeptidoleukotrienes). The oxazolone-induced contact sensitivity test inwhich mouse paw volume is measured is useful in detecting compoundswhich are immunostimulatory, but is of no known utility in detecting orsuggesting compounds which are inhibitors of the 5-lipoxygenase pathway.

SUMMARY OF THE INVENTION

This invention relates to a compound of the formula: ##STR2## wherein:

A is CH₂ CH₂ or CH₂ C(O)CH₂ ; and

R¹¹ and R₁₁₁ are both pyridyl or one of R¹¹

and R¹¹¹ is pyridyl and the other is monosubstituted phenyl wherein saidsubstituent is selected from halo; or a pharmaceutically acceptable saltthereof.

This invention relates to a pharmaceutical composition comprising aneffective, non-toxic 5-lipoxygenase pathway inhibiting amount of acompound of Formula (IA), or a pharmaceutically acceptable salt thereof,and a pharmaceutically acceptable carrier or diluent.

This invention also relates to a method of treating a 5-lipoxygenasepathway mediated disease in an animal in need thereof, provided thatsuch animal is in need of treatment of a 5-lipoxygenase pathway mediateddisease other than rheumatoid arthritis, which comprises administeringto such animal an effective, non-toxic 5-lipoxygenase pathway inhibitingamount of a compound of the formula ##STR3## wherein:

A is CH₂ CH₂ or CH₂ C(O)CH₂ ; and

R and R^(I) are independently selected from pyridyl

or monosubstituted phenyl wherein said substituent is selected fromhalo; or a pharmaceutically acceptable salt thereof.

DETAILED DESCRIPTION OF THE INVENTION

It will be apparent to one of skill in the art that all the compounds ofFormula (IA) are encompassed by the scope of Formula (I). All thecompounds of Formula (I) are useful in treating a 5-lipoxygenase pathwaymediated disease in an animal in need thereof by inhibiting the5-lipoxygenase pathway.

The preparation of compounds of Formula (I), other than those of Formula(IA) is disclosed in Hill, U.S. Pat. No. 4,188,397, issued Feb. 12,1980, the disclosure of which is hereby incorporated by reference.Preparation of compounds of Formula (IA) is accomplished by reacting theappropriate 4/5-(pyridyl)-5/4(pyridyl/halo substitutedphenyl)-[1H]-imidazole-2-thione [which can be prepared by the method ofLantos et al., J. Med. Chem, 27, 72-75 (1984], or alternately by themethod of Ciba Geigy, U.K. Patent Application GB No. 2,039,882, witheither 1-3-dihalopropanone or 1,2-dihaloethane in methanol.

Preferred compounds of Formula (I) include2,2'-[ethanediylbis(thio)]bis[4,5-bis(4-fluorophenyl)-1H-imidazole];2,2'-[1,3-propan-2-onediylbis(thio)]bis[4-(4-fluoro-phenyl)-5-(4-pyridyl)-1H-imidazole];and2,2'-[1,3-propan-2-onediylbis(thio)]bis[4,5-bis-(4-fluorophenyl)-1H-imidazole].

Pharmaceutically acceptable salts and their preparation are well knownto those skilled in pharmaceuticals. Pharmaceutically acceptable saltsof the compounds of Formula (I) which are useful in the presentinvention include, but are not limited to, hydrochloride, hydrobromidemaleate, fumarate, lactate, oxalate, methanesulfonate, ethanesulfonate,benzenesulfonate, tartrate, citrate, sulfate, phosphate and nitratesalts. Preparation of such salts for some Formula (I) compounds isdisclosed in Hill, U.S. Pat. No. 4,188,397.

It is known that some of the compounds of Formula (I) are useful fortreating cyclooxygenase product-mediated disease states. It has now beendiscovered that all of the compounds of Formula (I) are also useful fortreating disease states mediated by the 5-lipoxygenase pathway byinhibiting the 5-lipoxygenase pathway. The discovery that the compoundsof Formula (I) are inhibitors of the 5-lipoxygenase pathway is based onthe effects of the compounds of Formula (I) and on the production of5-lipoxygenase products by inflammatory cells in vitro in assays, someof which are described in the Examples. These data, together withprevious observations on the anti-edematous effects of some of thecompounds of Formula (I) in inflammatory lesions caused bycyclooxygenase-generated products, reveal that compounds of Formula (I)inhibit the 5-lipoxygenase pathway of arachidonic acid metabolism byshowing that they impair the production of 5-lipoxygenase products suchas leukotriene B₄ (di-HETE) and 5-HETE production by RBL-1 cells.

The pathophysiological role of arachidonic acid metabolites has been thefocus of recent intensive studies. In addition to the well-describedphlogistic activity (i.e. general inflammatory activity) ofprostaglandins, the more recent description of similar activity foreicosanoids has broadened the interest in these products as mediators ofinflammation [See, O'Flaherty, Lab. Invest., 47, 314-329 (1982)]. Thereported discovery of potent chemotactic and algesic activity for LTB₄[see, Smith, Gen. Pharmacol., 12, 211-216 (1981) and Levine et al.,Science, 225, 743-745 (1984)], together with known LTC₄ and LTD₄-mediated increase in capillary permeability [see, Simmons et al.,Biochem. Pharmacol., 32, 1353-1359 (1983), Veno et al., Prostaglandins,21, 637-647 (1981), and Camp et al., Br. J. Pharmacol., 80, 497-502(1983)], has led to their consideration as targets for phrmacologicalintervention in both the fluid and cellular phases of inflammatorydiseases.

The pharmacology of several inflammatory model systems has attested tothe effectiveness of corticosteroids in reducing the cellularinfiltration. These results, and the observation that corticosteroidsinhibit the generation of both cyclooxygenase and lipoxygenase products,suggest that such dual inhibitors may effectively reduce both the fluidand cellular phases of the inflammatory response since selectivecyclooxygenase inhibitors do not reliably inhibit cell influx intoinflammatory sites [See, Vinegar et al., Fed. Proc., 35, 2447-2456(1976), Higgs et al., Brit. Bull., 39, 265-270 (1983), and Higgs et al.,Prostaglandins, Leukotrienes and Medicine, 13, 89-92 (1984)]. Theobservations outlined above cogently argue that a dual inhibitor ofarachidonic acid metabolism would be a more effective antiinflammatoryagent than an inhibitor of cyclooxygenase only. Under optimalconditions, it is likely that an agent with preferential lipoxygenaseinhibitory activity would not share the ulcerogenic liability ofcyclooxygenase inhibitors or the toxicity of corticosteroids.

Recent clinical data also support the enthusiasm for 5-lipoxygenasepathway inhibitors of arachidonic acid metabolism in a variety ofinflammatory diseases in which granulocyte and/or monocyte infiltrationis prominent. The reported demonstration of elevated levels of LTB₄ inrheumatoid arthritic joint fluid [See, Davidson et al., Ann. Rheum.Dis., 42, 677-679 (1983)] also suggests a contributing role forarachidonic acid metabolites in rheumatoid arthritis. The recentlyreported preliminary observation of efficacy, including remission,reported with sulfasalazine treatment of rheumatoid arthritic patients[See Neumann et al., Brit. Med. J., 287, 1099-1102 (1983)] illustratesthe utility of inhibitors of the 5-lipoxygenase pathway in rheumatoidarthritis.

Sulfasalazine, which is used for treatment of ulcerative colitis, hasbeen reported to inhibit LTB₄ and 5-HETE production in vitro [See,Stenson et al., J. Clin. Invest., 69, 494-497 (1982)]. This observation,coupled with the fact that it has been reported that inflamedgastrointestinal mucosa from inflammatory bowel disease patients showedincreased production of LTB₄ [See, Sharon et al., Gastroenterol., 84,1306 (1983)], suggests that sulfasalazine can be effective by virtue ofinhibition of production of chemotactic eicosanoids (such as the5-lipoxygenase pathway product known as LTB₄). The observations serve tounderscore utility of inhibitors of the 5-lipoxygenase pathway ininflammatory bowel disease.

Another area of utility for inhibitors of the 5-lipoxygenase pathway isin the treatment of psoriasis. It was demonstrated that involvedpsoriatic skin had elevated levels of LTB₄ [See, Brain et al., Lancet,19, Feb. 19, 1983]. The promising effect of benoxaprofen on psoriasis[See, Allen et al., Brit. J. Dermatol., 109, 126-129 (1983)], a compoundwith in vitro lipoxygenase inhibitory activity on psoriasis, lendssupport to the concept that 5-lipoxygenase pathway inhibitors can beuseful in the treatment of psoriasis.

Lipoxygenase products have been identified in exudate fluids from goutypatients. This disorder is characterized by massive neutrophilinfiltration during the acute inflammatory phases of the disease. Sincea major 5-lipoxygenase product, LTB₄, is produced by neutrophils, itfollows that inhibition of the synthesis of LTB₄ can block anamplification mechanism in gout.

Another area in which inhibitors of the 5-lipoxygenase pathway can haveutility is in myocardial infarction. Studies in dogs with the dualinhibitor, BW755-C, demonstrated that the area of infarction followingcoronary occlusion was reduced, and such reduction was attributed toinhibition of leukocyte infiltration into the ischaemic tissue [See,Mollane et al., J. Pharmacol. Exp. Therap., 228, 510-522 (1984)].

Yet another area of utility for inhibitors of the 5-lipoxygenase pathwayis in the area of prevention of rejection of organ transplants. [See,e.g., Foegh et al., Adv. Prostaglandin, Thromboxane, and LeukotrieneResearch, 13, 209-217 (1983)].

Yet another utility for inhibitors of the 5-lipoxygenase pathway is inthe treatment of tissue trauma. [See, e.g., Denzliner et al., Science,230 (4723), 530-332 (1985)].

Furthermore, another area of utility for inhibitors of the5-lipoxygenase pathway is in the treatment of inflammatory reaction inthe central nervous system, including multiple sclerosis. [see, e.g.,Mackay et al., Clin. Exp. Immunol., 15, 471-482 (1973).

Another area of utility for inhibitors of the 5-lipoxygenase pathway isin the treatment of asthma. [see, e.g., Ford-Hutchinson, J. AllergyClin. Immunol., 74, 437-440 (1984).

This invention relates to a pharmaceutical composition comprising aneffective, 5-lipoxygenase pathway inhibiting amount of a compound ofFormula (IA), or a pharmaceutically acceptable salt thereof, and apharmaceutically acceptable carrier or diluent. As stated above, all ofthe compounds of Formula (IA) are within the scope of Formula (I).

The compounds of Formula (I) are administered in conventional dosageforms prepared by combining a compound of Formula (I) or a salt thereofin an amount sufficient to produce activity with a standardpharmaceutical carrier according to conventional procedures. Theseprocedures may involve mixing, granulating and compressing or dissolvingthe ingredients as appropriate to the desired preparation. The resultingpharmaceutical compositions which comprise an effective, 5-lipoxygenasepathway inhibiting amount of a compound of Formula (IA) are also objectsof this invention.

The pharmaceutical carrier employed may be, for example, either a solidor liquid. Exemplary of solid carriers are lactose, terra alba, sucrose,talc, gelatin, agar, pectin, acacia, magnesium stearate, stearic acidand the like. Exemplary of liquid carriers are syrup, peanut oil, oliveoil, water and the like. Similarly, the carrier or diluent may include atime delay material well known to the art, such as glyceryl monostearateor glyceryl distearate alone or with a wax.

A wide variety of pharmaceutical forms can be employed. Thus, if a solidcarrier is used, the preparation can be tableted, placed in a hardgelatin capsule in powder or pellet form or in the form of a troche orlozenge. The amount of solid carrier will vary widely but preferablywill be from about 25 mg to about 1 g. If a liquid carrier is used, thepreparation will be in the form of a syrup, emulsion, soft gelatincapsule, sterile injectable liquid such as an ampule or a nonaqueousliquid suspension.

To obtain a stable water soluble dose form, a pharmaceuticallyacceptable acid addition salt, preferably hydrochloride or sulfate, of acompound of Formula (I) is dissolved in an aqueous solution of anorganic or inorganic acid, such as a 0.3M solution of succinate acid or,preferably, citric acid. In addition to sulfate and hydrochloride,methanesulfonate, phosphate and hydrobromide are exemplary of otherwater soluble salts.

Preferably, each parenteral dosage unit will contain the activeingredient in an amount of from about 50 mg to about 500 mg.

The compounds of Formula (I) may be administered topically to a mammalin need of the inhibition of the 5-lipoxygenase pathway of arachidonicacid metabolism. Thus, the compounds of Formula (I) may be administeredtopically in the treatment or prophylaxis of inflammation in an animal,including man and other mammals and may be used in the relief ofrheumatoid spondylitis, osteoarthritis, gouty arthritis and otherarthritic conditions, inflamed joints, exzema, psoriasis or otherinflammatory skin conditions such as sunburn; inflammatory eyeconditions including conjunctivitis; pyresis, pain and other conditionsassociated with inflammation.

The amount of a compound of Formula (I) (hereinafter referred to as theactive ingredient) required for therapeutic effect on topicaladministration will, of course, vary with the compound chosen, thenature and severity of the inflammatory condition and the animalundergoing treatment, and is ultimately at the discretion of thephysician. A suitable anti-inflammatory dose of an active ingredient is1 ug to 500 mg of base for topical administration, the most preferreddosage being 1 ug to 1000 ug, for example, 5 to 25 ug administered twoor three times daily.

By topical administration is meant non-systemic administration andincludes the application of a compound of Formula (I) externally to theepidermis, to the buccal cavity and instillation of such a compound intothe ear, eye and nose, and where the compound does not significantlyenter the blood stream. By systemic administration is meant oral,intravenous, intraperitoneal and intramuscular administration.

While it is possible for an active ingredient to be administered aloneas the raw chemical, it is preferable to present it as a pharmaceuticalformulation. The active ingredient may comprise, for topicaladministration, from 0.001% to 10% w/w, e.g. from 1% to 2% by weight ofthe formulation although it may comprise as much as 10% w/w butpreferably not in excess of 5% w/w and more preferably from 0.1% to 1%w/w of the formulation.

The topical formulations of the present invention, both for veterinaryand for human medical use, comprise an active ingredient together withone or more acceptable carrier(s) thereof and optionally any othertherapeutic ingredient(s). The carrier(s) must be `acceptable` in thesense of being compatible with the other ingredients of the formulationand not deleterious to the recipient thereof.

Formulations suitable for topical administration include liquid orsemi-liquid preparations suitable for penetration through the skin tothe site of inflammation such as: liniments, lotions, creams, ointmentsor pastes, and drops suitable for administration to the eye, ear ornose.

Drops according to the present invention may comprise sterile aqueous oroily solutions or suspensions and may be prepared by dissolving theactive ingredient in a suitable aqueous solution of a bactericidaland/or fungicidal agent and/or any other suitable preservative, andpreferably including a surface active agent. The resulting solution maythen be clarified by filtration, transferred to a suitable containerwhich is then sealed and sterilized by autoclaving or maintaining at98°-100° C. for half an hour. Alternatively, the solution may besterilized by filtration and transferred to the container by an aseptictechnique. Examples of bactericidal and fungicidal agents suitable forinclusion in the drops are phenylmercuric nitrate or acetate (0.002%),benzalkonium chloride (0.01%) and chlorhexidine acetate (0.01%).Suitable solvents for the preparation of an oily solution includeglycerol, diluted alcohol and propylene glycol.

Lotions according to the present invention include those suitable forapplication to the skin or eye. An eye lotion may comprise a sterileaqueous solution optionally containing a bactericide and may be preparedby methods similar to those for the preparation of drops. Lotions orliniments for application to the skin may also include an agent tohasten drying and to cool the skin, such as an alcohol or acetone,and/or a moisturizer such as glycerol or an oil such as castor oil orarachis oil.

Creams, ointments or pastes according to the present invention aresemi-solid formulations of the active ingredient for externalapplication. They may be made by mixing the active ingredient infinely-divided or powdered form, alone or in solution or suspension inan aqueous or non-aqueous fluid, with the aid of suitable machinery,with a greasy or non-greasy basis. The basis may comprise hydrocarbonssuch as hard, soft or liquid paraffin, glycerol, beeswax, a metallicsoap; a mucilage; an oil of natural origin such as almond, corn,arachis, castor or olive oil; wool fat or its derivatives, or a fattyacid such as steric or oleic acid together with an alcohol such asprolylene glycol or macrogols. The formulation may incorporate anysuitable surface active agent such as an anionic, cationic or non-ionicsulfactant such as sorbitan esters or polyoxyethylene derivativesthereof. Suspending agents such as natural gums, cellulose derivativesor inorganic materials such as silicaceous silicas, and otheringredients such as lanolin, may also be included.

The compounds of Formula (I) may also be administered by inhalation. By"inhalation" is meant intranasal and oral inhalation administration.Appropriate dosage forms for such administration, such as an aerosolformulation or a metered dose inhaler, may be prepared by conventionaltechniques. The preferred daily dosage amount of a compound of Formula(I) administered by inhalation is from about 10 mg to about 200 mg perday.

This invention relates to a method of treating a disease state mediatedby the 5-lipoxygenase pathway in an animal in need thereof, includinghumans and other mammals, provided that such animal is in need oftreatment of a 5-lipoxygenase pathway mediated disease other than,rheumatoid arthritis, which comprises administering to such animal aneffective, 5-lipoxygenase pathway inhibiting amount of a Formula (I)compound or a pharmaceutically acceptable salt thereof. By "treating" ismeant either prophylactic or therapeutic therapy. By "mediated" is meantcaused by or exacerbated by. The formula (I) compound is administered toan animal in need of inhibition of the 5-lipoxygenase pathway in anamount sufficient to inhibit the 5-lipoxygenase pathway. Such Formula(I) compound can be administered to such animal in a conventional dosageform prepared by combining the Formula (I) compound with a conventionalpharmaceutically acceptable carrier or diluent according to knowntechniques. It will be recognized by one of skill in the art that theform and character of the pharmaceutically acceptable carrier or diluentis dictated by the amount of active ingredient with which it is to becombined, the route of administration and other well-known variables.The route of administration may be parenteral, by inhalation or topical.The term parenteral as used herein includes intravenous, intramuscular,subcutaneous, intrarectal, intravaginal or intraperitonealadministration. The subcutaneous and intramuscular forms of parenteraladministration are generally preferred. The daily dosage regimen forparenteral administration will preferably be from about 100 mg to about1.5 g per day. The daily dosage regimen for topical administration willpreferably be from about 2 mg to about 10 mg. per site ofadministration.

It will be recognized by one of skill in the art that the optimalquantity and spacing of individual dosages of the Formula (I) compoundwill be determined by the nature and extent of the condition beingtreated, the form, route and site of administration, and the particularanimal being treated, and that such optimums can be determined byconventional techniques. It will also be appreciated by one of skill inthe art that the optimal course of treatment, i.e., the number of dosesof the Formula (I) compound given per day for a defined number of days,can be ascertained by those skilled in the art using conventional courseof treatment determination tests.

EXAMPLES

Without further elaboration, it is believed that one skilled in the artcan, using the preceding description, utilize the present invention toits fullest extent. The following examples are, therefore, to beconstrued as merely illustrative and not a limitation of the scope ofthe present invention in any way.

SYNTHESIS EXAMPLES Example 12-2'-[1,3-propan-2-onediylbis(thio)]bis[5-(4-pyridyl)-4-(4-fluorophenyl)-1H-imidazole]

A suspension of 4-(4-pyridyl)-5-(4-fluorophenyl)-[1H]-imidazole-2-thione(5.0 g, 0.0184 mol), prepared by the method of Lantos et al., J. Med.Chem., 27, 72-75 (1984), and 1,3-dichloropropanone (1.15 g, 0.00906 mol)in ethanol (30 ml) was refluxed for 90 minutes, and then the mixture wascooled. The resulting precipitate was filtered, washed with ether andrecrystallized from ethanol to give the title compound as thedihydrochloride salt (hydrate), melting point (mp) 251°-252° C. (dec).

C₃₁ H₂₂ F₂ N₆ OS₂ 2HCl0.75H₂ O

Calculated: 54.51%C; 3.76%H; 12.30%N. Found: 54.46%C; 3.82%H; 12.46%N.

UTILITY EXAMPLES

In the following Examples male Balb/c mice (20-28 g) were used. All micewere obtained from Charles River Breeding Laboratories, Kingston, N.Y.Within a single experiment, mice were sex and age matched.

In the following examples, reagents used were employed as follows:

Compounds of Formula (I), indomethacin, naproxen, and ibuprofen wereeach used as the free base. The compounds were homogenized in 0.5%tragacanth. Compounds were administered by gavage at the indicated dosein a final volume of 10 ml/kg.

For in vitro experiments, compounds were dissolved at appropriateconcentrations in ethanol (final concentration 1.0%) and then diluted tofinal concentrations using the buffers indicated in the text.

I. METHODS Arachidonic Acid-Induced Mouse Ear Inflammation

Arachidonic acid in acetone (2 mg/20 μl) was applied to the innersurface of the left ear. The thickness of both ears was then measuredwith a dial micrometer one hour after treatment, and the data wereexpressed as the change in thickness (10⁻³ cm) between treated anduntreated ears.

Test compounds were given orally in 0.5% tragacanth at the timesindicated in the test prior to the topical application of arachidonicacid.

Parenteral administration of compound was accomplished by subcutaneousinjection of solution as indicated.

Assay of 5-Lipoxygenase Activity

The activities of these enzymes in extracts of RBL-1 cells were assayedusing the method of Jakschik and Lee, Nature, 287, 51-52 (1980). RBL-1cells were obtained from the American Type Culture Collection (#CRL1378) and were grown at 37° C. (5% CO₂ in air) in spinner culture in MEMsupplemented with 10% heat inactivated fetal calf serum. Harvested cellswere washed with 50 mM sodium phosphate buffer, pH 7.0, containing 1 mMEDTA and 0.1% gelatin, resuspended in fresh buffer (5×10⁷ cells/ml) anddisrupted by nitrogen cavitation using the Parr bomb at 750 psi for 10min. The broken cell extract was then centrifuged at 10,000×g for 20minutes (min) and the supernatant centrifuge at 100,000×g for 60 min.Aliquots (0.25 mls) of the supernatant were preincubated with or withoutdrugs for 10 min, after which 12 mM CaCl₂ was added and the reaction wasinitiated with 2.5 ml of 2.5 μM arachidonic acid-1-¹⁴ C (finalconcentration was 25 μM; specific activity 20,000 dpm/nmole). Afterincubation for 5 min at 37° C., the reaction was terminated by additionof 2 volumes (0.5 ml) ice cold acetone and the sample was allowed todeproteinize on ice for 10 min prior to centrifugation at 1,000×g for 10min. The deproteinized supernatant was adjusted to pH 3.5 with 2N formicacid and extracted with 2 volumes of ice cold ethyl acetate. Theextracted samples were dried under argon, redissolved in ethyl acetateand applied to Whatman LK5D thin layer chromatography (TLC) plates whichwere developed using the A-9 solvent system [organic phase of ethylacetate: 2,2,5-trimethylpentane:acetic acid:water (110:50:20:10)]described by Hamberg and Samuelsson, J. Biol. Chem., 241, 257-263(1966). Arachidonic acid, 5-HETE, LTB₄ and PGD₂ were quantified with aBerthold LB 2832 autoscanner.

Under these conditions, only the 5-lipoxygenase pathway metabolites weredetectable. The 5-HETE and di-HETEs were formed at a linear rate, andsubstantial amounts of the arachidonic acid-1-¹⁴ C substrate wereutilized.

Drug-induced effects on enzyme activities are described as theconcentration of drug causing a 50% inhibition of metabolite synthesis(IC₅₀).

II. RESULTS The Effect of Compounds of Formula (I) on ArachidonicAcid-induced Inflammation

Compounds of Formula (I) dosed orally produce insignificant inhibitionof the edematous response normally seen 1 hour after the application of2 mg of arachidonic acid to the ear. The cyclooxygenase inhibitors,indomethacin (10 mg/kg, p.o.), ibuprofen (250 mg/kg, p.o.) and naproxen(100 mg/kg, p.o.) also did not exhibit detectable antiinflammatoryactivity in this assay, despite use at near maximally tolerated doses.

The Effect of Compounds of Formula (I) on Arachidonic Acid Metabolism

Experiments using a soluble extract preparation of RBL-1 cellscontaining only lipoxygenase activity confirmed the inhibitory effectsof compounds of Formula (I) on LTB₄ production (Table II) and 5-HETEproduct (Table III). The data in Table II shows that compounds ofFormula (I) are significant inhibitors of the 5-lipoxygenase pathway asconfirmed by their inhibition of LTB₄, a 5-lipoxygenase pathway product.The data in Table III also shows that compounds of Formula (I) aresignificant inhibitors of the 5-lipoxygenase pathway as confirmed bytheir inhibition of 5-HETE, a 5-lipoxygenase pathway product. Thus,although the compounds of Formula (I) are insignificant inhibitors ofarachidonic acid-induced inflammation when dosed orally (Table 1), thedata of Table II and Table III confirm that such compounds are indeedinhibitors of the 5-lipoxygenase pathway.

                  TABLE I                                                         ______________________________________                                        The Effect of Compounds of Formula (I) on                                     Arachidonic Acid Induced Ear Swelling                                          ##STR4##                 Formula (I)                                                                                %                                                                             Inhibi-                                Com-                                   tion.sup. (a,b)                        pound                                  of Ear                                 Num-                                   Swell-                                 ber   R           R.sup.1     A        ing                                    ______________________________________                                        1     4-fluorophenyl                                                                            4-fluorophenyl                                                                            CH.sub.2 CH.sub.2                                                                      N                                                                             (p.o.)                                 2     4-pyridyl   4-fluorophenyl                                                                            CH.sub.2 C(O)CH.sub.2                                                                  NS                                                                            (p.o.)                                 3     4-fluorophenyl                                                                            4-fluorophenyl                                                                            CH.sub.2 C(O)CH.sub.2                                                                  NT                                     ______________________________________                                         .sup.(a) screened at 50 mg/kg s.c. or i.p. unless indicated as oral dosin     (p.o.).                                                                       .sup.(b) * = p < .05, ** = p < .01, *** = p < .001, NS = not significant,      NT = not tested.                                                        

                  TABLE II                                                        ______________________________________                                        The Effect of Compounds of Formula (I) on                                     5-Lipoxygenase Activity                                                        ##STR5##                 Formula (I)                                         Com-                                                                          pound                                  LIB.sub. 4                             Num-                                   IC.sub.50                              ber   R           R.sup.1     A        (μM)                                ______________________________________                                        1     4-fluorophenyl                                                                            4-fluorophenyl                                                                            CH.sub.2 CH.sub.2                                                                      3.3                                    2     4-pyridyl   4-fluorophenyl                                                                            CH.sub.2 C(O)CH.sub.2                                                                  0.65                                   3     4-fluorophenyl                                                                            4-fluorophenyl                                                                            CH.sub.2 C(O)CH.sub.2                                                                  1.6                                    ______________________________________                                         .sup.(a) IC.sub.50 determined on LTB.sub.4 production by RBL1 high speed      supernatant.                                                             

                                      TABLE III                                   __________________________________________________________________________    The Effect of Compounds of Formula (I) on                                     5-Lipoxygenase Activity (5-HETE Production)                                    ##STR6##               Formula (I)                                                                            5-HETE.sup.(a)                               Compound Number                                                                         R       R.sup.1 A      IC.sub.50 (μM)                            __________________________________________________________________________    1         4-fluorophenyl                                                                        4-fluorophenyl                                                                        CH.sub.2 CH.sub.2                                                                    18.5                                         2         4-pyridyl                                                                             4-fluorophenyl                                                                        CH.sub.2 C(O)CH.sub.2                                                                1.7                                          3         4-fluorophenyl                                                                        4-fluorophenyl                                                                        CH.sub.2 C(O)CH.sub.2                                                                5.3                                          __________________________________________________________________________     .sup.(a) IC.sub.50 determined on 5HETE production by RBL1 high speed          supernatant.                                                             

EXAMPLE A Injectable Parenteral Composition

A pharmaceutical composition of this invention in a form suitable foradministration by injection is prepared by stirring 10% by weight of acompound of Formula (IA) in 10% by volume propylene glycol and water.

EXAMPLE B Ointment Composition

Compound of Formula (IA) 1.0 g

White soft paraffin to 100.0 g

The compound of Formula (IA) is dispersed in a small volume of thevehicle and this dispersion is gradually incorporated into the bulk toproduce a smooth, homogeneous product which is filled into collapsiblemetal tubes.

EXAMPLE C Topical Cream Composition

Compound of Formula (IA) 1.0 g

Polawax GP 200 20.0 g

Lanolin Anhydrous 2.0 g

White Beeswax 2.5 g

Methyl hydroxybenzoate 0.1 g

Distilled Water to 100.0 g

The polawax, beeswax and lanolin are heated together at 60° C. and asolution of methyl hydroxybenzoate is added. Homogenization is achievedusing high speed stirring and the temperature is allowed to fall to 50°C. The compound of Formula (IA) is added and dispersed throughout, andthe composition is allowed to cool with slow speed stirring.

EXAMPLE D Topical Lotion Composition

Compound of Formula (IA) 1.0 g

Sorbitan Monolaurate 0.6 g

Polysorbate 20 0.6 g

Cetostearyl Alcohol 1.5 g

Glycerin 6.0 g

Methyl hydroxybenzoate 0.2 g

Purified Water B.P. to 100.00 ml

The methyl hydroxybenzoate and glycerin are dissolved in 70 ml of thewater at 75°. The sorbitan monolaurate, polysorbate 20 and cetostearylalcohol are melted together at 75° C. and added to the aqueous solution.The resulting emulsion is homogenized, allowed to cool with continuousstirring and the compound of Formula (IA) is added as a suspension inthe remaining water. The whole suspension is stirred until homogenized.

EXAMPLE E Eye Drop Composition

Compound of Formula (IA) 0.5 g

Methyl Hydroxybenzoate 0.01 g

Propyl Hydroxybenzoate 0.04 g

Purified Water B.P. to 100.00 ml

The methyl and propyl hydroxybenzoates are dissolved in 70 ml purifiedwater at 75° C. and the resulting solution is allowed to cool. Thecompound of Formula (IA) is then added, and the solution is made up to100 ml with purified water. The solution is sterilized by filtrationthrough a membrane filter (0.22 mu m pore size) and packed asepticallyinto suitable sterile containers.

EXAMPLE F Composition for Administration by Inhalation

For an aerosol container with a capacity of 15-20 ml: mix 10 mg of acompound of Formula (IA) with 0.1-0.2% of a lubricating agent, such asSpan 85 or oleic acid, and disperse such mixture in a propellane (c.a.),such as freon, preferably a combination of freon 114 and freon 12, andput into an appropriate aerosol container adapted for either intranasalor oral inhalation administration.

EXAMPLE G Composition for Administration by Inhalation

For an aerosol container with a capacity of 15-20 ml: Dissolve 10 mg ofa compound of Formula (IA) in ethanol (6-8 ml) and disperse such in apropellant (c.a.), such as freon, preferably a combination of freon 114and freon 12, and put into an appropriate aerosol container adapted foreither intranasal or oral inhalation administration.

What is claimed is:
 1. A method of treating a disease mediated by the5-lipoxygenase pathway in an animal in need thereof, provided suchanimal is in need of treatment of one of the 5-lipoxygenase pathwaymediated diseases of inflammatory bowel disease, psoriasis, gout,myocardial infarction, organ transplant rejection, tissue trauma,multiple sclerosis, asthma, rheumatoid spondylitis, osteoarthritis,gouty arthritis, inflamed joints, exzema, conjunctivitis or pyresis,which comprises administering parenterally, topically or by inhalationto such animal an effective, non-toxic 5-lipoxygenase pathway inhibitingamount of a compound of the formula ##STR7## wherein: A is CH₂ CH₂ orCH₂ C(O)CH₂ ; andR and R¹ are pyridyl or one of R and R¹ is pyridyl andthe other is monosubstituted phenyl wherein said substituent is selectedfrom halo; or a pharmaceutically acceptable salt thereof.
 2. The methodof claim 1 wherein A is CH₂ C(O)CH₂, R is 4-pyridyl and R¹ is4-fluorophenyl.
 3. The method of claim 1 wherein the administration isparenteral and the amount of compound administered is selected fromabout 50 mg to about 500 mg.
 4. The method of claim 1 wherein the amountof compound administered per day is from about 50 mg to about 1000 g. 5.The method of claim 1 wherein the compound is administered byinhalation.
 6. The method of claim 5 wherein the amount of compoundadministered is from about 10 mg to about 200 mg per day.
 7. The methodof claim 1 wherein the compound is administered topically.
 8. The methodof claim 7 wherein the amount of compound administered per dose is 1 μgto 1000 μg.